Project Summary: Primary sleep disorders are among the most common medical conditions, and clinical data show women are far more likely to experience sleep disorders over their lifespan. This increased risk emerges at puberty and has been associated with fluctuations in ovarian steroids, particularly estrogens, suggesting that gonadal steroids and biological sex are significant risk factors for sleep disruptions. Despite a growing understanding of sleep regulatory mechanisms, how estrogens influence the sleep circuitry is poorly understood. Historically, male rodents have served as the cornerstone for elucidating the neural circuitries governing sleep. Unfortunately, this has resulted in a significant gap in our understanding of how estrogens modulate these circuits in females. The female rodent offers the opportunity to probe the sleep circuitry to elucidate the mechanisms by which ovarian steroids modulate sleep, as sleep patterns in the female rat are exquisitely sensitive to natural fluctuations in ovarian steroids such as estradiol (E2). Using adult female Sprague-Dawley rats, our group has made inroads into understanding estrogenic influences over normal wake and sleep patterns. Our studies consistently demonstrate that sleep time is significantly reduced when endogenous ovarian steroids or exogenous E2 are elevated in females but not males. This effect is mediated through the inhibition of sleep-active neurons in the median preoptic nucleus (MnPN). Moreover, we have demonstrated that E2 is capable of marked suppression of sleep under sleep deprivation, when homeostatic sleep need, also known as sleep pressure, is increased. Taken together with our previous findings, this observation suggests that E2 may dissipate the homeostatic need for sleep. Furthermore, preliminary data suggest that E2 attenuates the action of adenosine signaling at the sleep-promoting A2A receptor, resulting in reduced sleep duration. This finding is significant because adenosine is a known mediator of sleep pressure, with established actions at the MnPN and a closely related (but non-E2 sensitive) nucleus, the ventral lateral preoptic (VLPO). These data suggest that there is an interplay between E2 and adenosine which modulates the ability of adenosine to generate sleep pressure. However, the specific mechanism for this interplay remains undetermined. This project will test the hypothesis that estrogenic modulation of sleep pressure requires the attenuation of adenosine signaling efficacy. We will test this hypothesis by (1) determining if estradiol reduces the homeostatic need for sleep through both behavioral experiments and molecular assays of adenosine content, synthesis, and clearance, and (2) determining if estradiol attenuates neuronal activity of sleep active MnPN neurons, through inhibiting the cellular action of adenosine in the sleep circuitry, through experiments in both receptor pharmacology and neuronal activity. Understanding interactions of E2 and adenosine may illuminate ties between hormonal content and sleep, and provide opportunities for sleep disorder treatments that take female sex and hormonal state into account.